A Critical Analysis of Atoh7 (Math5) mRNA Splicing in the Developing Mouse Retina

The Math5 (Atoh7) gene is transiently expressed during retinogenesis by progenitors exiting mitosis, and is essential for ganglion cell (RGC) development. Math5 contains a single exon, and its 1.7 kb mRNA encodes a 149-aa polypeptide. Mouse Math5 mutants have essentially no RGCs or optic nerves. Given the importance of this gene in retinal development, we thoroughly investigated the possibility of Math5 mRNA splicing by Northern blot, 3′RACE, RNase protection assays, and RT-PCR, using RNAs extracted from embryonic eyes and adult cerebellum, or transcribed in vitro from cDNA clones. Because Math5 mRNA contains an elevated G+C content, we used graded concentrations of betaine, an isostabilizing agent that disrupts secondary structure. Although ∼10% of cerebellar Math5 RNAs are spliced, truncating the polypeptide, our results show few, if any, spliced Math5 transcripts exist in the developing retina (<1%). Rare deleted cDNAs do arise via RT-mediated RNA template switching in vitro, and are selectively amplified during PCR. These data differ starkly from a recent study (Kanadia and Cepko 2010), which concluded that the vast majority of Math5 and other bHLH transcripts are spliced to generate noncoding RNAs. Our findings clarify the architecture of the Math5 gene and its mechanism of action. These results have implications for all members of the bHLH gene family, for any gene that is alternatively spliced, and for the interpretation of all RT-PCR experiments

Assessing patterns of hybridization between North Atlantic eels using diagnostic single-nucleotide polymorphisms

The two North Atlantic eel species, the European eel (Anguilla anguilla) and the American eel (Anguilla rostrata), spawn in partial sympatry in the Sargasso Sea, providing ample opportunity to interbreed. In this study, we used a RAD (Restriction site Associated DNA) sequencing approach to identify species-specific diagnostic single-nucleotide polymorphisms (SNPs) and design a low-density array that combined with screening of a diagnostic mitochondrial DNA marker. Eels from Iceland (N=159) and from the neighboring Faroe Islands (N=29) were genotyped, along with 94 larvae (49 European and 45 American eel) collected in the Sargasso Sea. Our SNP survey showed that the majority of Icelandic eels are pure European eels but there is also an important contribution of individuals of admixed ancestry (10.7%). Although most of the hybrids were identified as F1 hybrids from European eel female × American eel male crosses, backcrosses were also detected, including a first-generation backcross (F1 hybrid × pure European eel) and three individuals identified as second-generation backcrosses originating from American eel × F1 hybrid backcrosses interbreeding with pure European eels. In comparison, no hybrids were observed in the Faroe Islands, the closest bodies of land to Iceland. It is possible that hybrids show an intermediate migratory behaviour between the two parental species that ultimately brings hybrid larvae to the shores of Iceland, situated roughly halfway between the Sargasso Sea and Europe. Only two hybrids were observed among Sargasso Sea larvae, both backcrosses, but no F1 hybrids, that points to temporal variation in the occurrence of hybridization

Reactivation of the inactive X chromosome in development and reprogramming

In mammals, one of the two X chromosomes of female cells is inactivated for dosage compensation between the sexes. X chromosome inactivation is initiated in early embryos by the noncoding Xist RNA. Subsequent chromatin modifications on the inactive X chromosome (Xi) lead to a remarkable stability of gene repression in somatic cell lineages. In mice, reactivation of genes on the Xi accompanies the establishment of pluripotent cells of the female blastocyst and the development of primordial germ cells. Xi reactivation also occurs when pluripotency is established during the reprogramming of somatic cells to induced pluripotent stem cells. The mechanism of Xi reactivation has attracted increasing interest for studying changes in epigenetic patterns and for improving methods of cell reprogramming. Here, we review recent advances in the understanding of Xi reactivation during development and reprogramming and illustrate potential clinical applications

Genomics and the origin of species

Speciation is a fundamental evolutionary process, the knowledge of which is crucial for understanding the origins of biodiversity. Genomic approaches are an increasingly important aspect of this research field. We review current understanding of genome-wide effects of accumulating reproductive isolation and of genomic properties that influence the process of speciation. Building on this work, we identify emergent trends and gaps in our understanding, propose new approaches to more fully integrate genomics into speciation research, translate speciation theory into hypotheses that are testable using genomic tools and provide an integrative definition of the field of speciation genomics